Neuropathology of patients with COVID-19 in Germany: a post-mortem case series

Abstract

Background: Prominent clinical symptoms of COVID-19 include CNS manifestations. However, it is unclear whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, gains access to the CNS and whether it causes neuropathological changes. We investigated the brain tissue of patients who died from COVID-19 for glial responses, inflammatory changes, and the presence of SARS-CoV-2 in the CNS.

Methods: In this post-mortem case series, we investigated the neuropathological features in the brains of patients who died between March 13 and April 24, 2020, in Hamburg, Germany. Inclusion criteria comprised a positive test for SARS-CoV-2 by quantitative RT-PCR (qRT-PCR) and availability of adequate samples. We did a neuropathological workup including histological staining and immunohistochemical staining for activated astrocytes, activated microglia, and cytotoxic T lymphocytes in the olfactory bulb, basal ganglia, brainstem, and cerebellum. Additionally, we investigated the presence and localisation of SARS-CoV-2 by qRT-PCR and by immunohistochemistry in selected patients and brain regions.

Findings: 43 patients were included in our study. Patients died in hospitals, nursing homes, or at home, and were aged between 51 years and 94 years (median 76 years [IQR 70-86]). We detected fresh territorial ischaemic lesions in six (14%) patients. 37 (86%) patients had astrogliosis in all assessed regions. Activation of microglia and infiltration by cytotoxic T lymphocytes was most pronounced in the brainstem and cerebellum, and meningeal cytotoxic T lymphocyte infiltration was seen in 34 (79%) patients. SARS-CoV-2 could be detected in the brains of 21 (53%) of 40 examined patients, with SARS-CoV-2 viral proteins found in cranial nerves originating from the lower brainstem and in isolated cells of the brainstem. The presence of SARS-CoV-2 in the CNS was not associated with the severity of neuropathological changes.

Interpretation: In general, neuropathological changes in patients with COVID-19 seem to be mild, with pronounced neuroinflammatory changes in the brainstem being the most common finding. There was no evidence for CNS damage directly caused by SARS-CoV-2. The generalisability of these findings needs to be validated in future studies as the number of cases and availability of clinical data were low and no age-matched and sex-matched controls were included.

Funding: German Research Foundation, Federal State of Hamburg, EU (eRARE), German Center for Infection Research (DZIF).

Figure 1

Common neuropathological findings in the brains of patients who died from COVID-19 An overview of each brain region with haematoxylin and eosin staining is shown in the first column. Immunohistochemical staining for the astrocytic marker GFAP showed variable degrees of reactive astrogliosis. Immunohistochemical staining for the microglia marker HLA-DR showed reactive activation of the microglia with occasional microglial nodules in the medulla oblongata and cerebellum (green arrows). Staining for the cytotoxic T lymphocyte marker CD8 (brown) revealed perivascular and parenchymal infiltration with CD8-positive cells. GFAP=glial fibrillary acidic protein.

Figure 2

Concomitant activation of the adaptive and innate immune systems in the brain of one patient (case 2) who died from COVID-19 Representative images of double-chromogenic immunohistochemical labelling for IBA1 (brown) and CD8 (pink), as well as immunohistochemical staining for CD68 (brown), and TMEM119 (brown) at different CNS interfaces in the upper medulla oblongata. Counterstaining was done with haematoxylin (blue). Scale bars represent 100 μm (10 μm in the inset images). Arrows indicate CD8-positive T cells.

Figure 3

In-silico analysis of the distribution of genes relevant to severe acute respiratory syndrome coronavirus 2 in the CNS Human temporal lobe cell type-specific expression of TMPRSS2, TMPRSS4, CTSL, TPCN2, NRP1, and ACE2. The heatmap shows the per-gene normalised mean expression across cell types (expression sums to 1 across the cell types). OPC=oligodendrocyte precursor cell.

Figure 4

Neuropathological findings and SARS-CoV-2 viral loads in studied patients (n=43) Cases are arranged from left to right on the basis of the presence and quantity of SARS-CoV-2 in the brain. F=female. FFPE=formalin-fixed paraffin-embedded. HPF=high-power field. IHC=immunohistochemistry. M=male. P=parenchymal. PV=perivascular. qPCR=quantitative PCR. SARS-CoV-2=severe acute respiratory syndrome coronavirus 2. *Values shown for positive cases represent number of copies of SARS-CoV-2 RNA (× 10³/mL); detection was done in the frontal lobe in cryopreserved specimens and in the upper medulla oblongata in FFPE specimens.

Figure 5

Distribution of SARS-CoV-2 within the CNS Representative images of viral protein-positive cells (green arrows) in the medulla oblongata detected by anti-nucleocapsid protein antibody (A) or anti-spike protein antibody (B). (C) SARS-CoV-2 nucleoprotein (brown staining) could also be detected in subsets of cranial nerves originating from the lower brainstem. SARS-CoV-2=severe acute respiratory syndrome coronavirus 2.